Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V007843 | pDSK519 | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pDSK519
- Antibiotic Resistance:
- Kanamycin
- Length:
- 8699 bp
- Type:
- Cloning vector
- Replication origin:
- RSF1010 oriV
- Source/Author:
- Keen NT, Tamaki S, Kobayashi D, Trollinger D.
pDSK519 vector Map
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5. Store the plasmid at -20 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pDSK519 vector Sequence
LOCUS 40924_15590 8699 bp DNA circular SYN 17-DEC-2018
DEFINITION Cloning vector pDSK519, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8699)
AUTHORS Keen NT, Tamaki S, Kobayashi D, Trollinger D.
TITLE Improved broad-host-range plasmids for DNA cloning in gram-negative
bacteria
JOURNAL Gene 70 (1), 191-197 (1988)
PUBMED 2853689
REFERENCE 2 (bases 1 to 8699)
AUTHORS Korte HL, Wall JD.
TITLE Direct Submission
JOURNAL Submitted (29-NOV-2011) Biochemistry, University of Missouri, 117
Schweitzer Hall, Columbia, MO 65211, USA
REFERENCE 3 (bases 1 to 8699)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 8699)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Gene";
date: "1988"; volume: "70"; issue: "1"; pages: "191-197"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(29-NOV-2011) Biochemistry, University of Missouri, 117 Schweitzer
Hall, Columbia, MO 65211, USA"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..8699
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 9..25
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
misc_feature complement(38..94)
/label=MCS
/note="pUC18/19 multiple cloning site"
primer_bind complement(95..111)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
repeat_region 631..991
/label=Tn903 inverted repeat
/note="Tn903 inverted repeat"
CDS 1096..1908
/codon_start=1
/label=KanR
/note="aminoglycoside phosphotransferase"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
repeat_region 1972..2332
/label=Tn903 inverted repeat
/note="Tn903 inverted repeat"
rep_origin complement(2725..3119)
/direction=LEFT
/label=RSF1010 oriV
/note="replication origin of the broad-host-range plasmid
RSF1010; requires the RSF1010 RepA/B/C proteins for
replication (Scholz et al., 1989)"
CDS complement(3146..3430)
/codon_start=1
/gene="mobC"
/product="MobC"
/label=mobC
/note="mobilization protein C"
/protein_id="AFG33234.1"
/translation="MVKGSNKAADRLAKLEEQRARINAEIQRVRAREQQQERKNETRRK
VLVGAMILAKVNSSEWPEDRLMAAMDAYLERDHDRALFGLPPRQKDEPG"
gene complement(3146..3430)
/gene="mobC"
/label=mobC
oriT 3461..3548
/label=RSF1010 oriT
/note="origin of transfer of the broad-host-range plasmid
RSF1010 (Scholz et al., 1989)"
CDS 4377..4790
/codon_start=1
/gene="mobB"
/product="MobB"
/label=mobB
/note="mobilization protein B"
/protein_id="AFG33235.1"
/translation="MNAIDRVKKSRGINELAEQIEPLAQSMATLADEARQVMSQTQQAS
EAQAAEWLKAQRQTGAAWVELAKELREVAAEVSSAAQSARSASRGWHWKLWLTVMLASM
MPTVVLLIASLLLLDLTPLTTEDGSIWLRLVAR"
gene 4377..4790
/gene="mobB"
/label=mobB
CDS 4787..5755
/codon_start=1
/label=RSF1010 RepB
/note="replication protein B of the broad-host-range
plasmid RSF1010 (Scholz et al., 1989)"
/translation="MKNDRTLQAIGRQLKAMGCERFDIGVRDATTGQMMNREWSAAEVL
QNTPWLKRMNAQGNDVYIRPAEQERHGLVLVDDLSEFDLDDMKAEGREPALVVETSPKN
YQAWVKVADAAGGELRGQIARTLASEYDADPASADSRHYGRLAGFTNRKDKHTTRAGYQ
PWVLLRESKGKTATAGPALVQQAGQQIEQAQRQQEKARRLASLELPERQLSRHRRTALD
EYRSEMAGLVKRFGDDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
CDS 6269..7105
/codon_start=1
/label=RSF1010 RepA
/note="replication protein A of the broad-host-range
plasmid RSF1010 (Scholz et al., 1989)"
/translation="MATHKPINILEAFAAAPPPLDYVLPNMVAGTVGALVSPGGAGKSM
LALQLAAQIAGGPDLLEVGELPTGPVIYLPAEDPPTAIHHRLHALGAHLSAEERQAVAD
GLLIQPLIGSLPNIMAPEWFDGLKRAAEGRRLMVLDTLRRFHIEEENASGPMAQVIGRM
EAIAADTGCSIVFLHHASKGAAMMGAGDQQQASRGSSVLVDNIRWQSYLSSMTSAEAEE
WGVDDDQRRFFVRFGVSKANYGAPFADRWFRRHDGGVLKPAVLERQRKSKGVPRGEA"
CDS 7095..7943
/codon_start=1
/label=RSF1010 RepC
/note="replication protein C of the broad-host-range
plasmid RSF1010 (Scholz et al., 1989)"
/translation="VVKPKNKHSLSHVRHDPAHCLAPGLFRALKRGERKRSKLDVTYDY
GDGKRIEFSGPEPLGADDLRILQGLVAMAGPNGLVLGPEPKTEGGRQLRLFLEPKWEAV
TADAMVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
TGKASIDTLCGYVWPSEASGSTMRKRRQRVREALPELVALGWTVTEFAAGKYDITRPKA
AG"
protein_bind 8610..8631
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 8646..8676
/label=lac promoter
/note="promoter for the E. coli lac operon"